The chorion of Aedes aegypti eggs undergoes a hardening process following oviposition and individual chorion proteins become insoluble thereafter. Our previous studies determined that peroxidase-catalyzed chorion protein crosslinking and phenoloxidase-mediated chorion melanization are primarily responsible for the formation of a hardened, desiccation resistant chorion in A. aegypti eggs. To gain further understanding of peroxidase- and phenoloxidase-catalyzed biochemical processes during chorion hardening, we analyzed chorion proteins, identified three low molecular weight major endochorion proteins that together constituted more than 70% of the total amount of endochorion proteins, and assessed their insolubilization in relation to phenoloxidase- and peroxidase-catalyzed reactions under different conditions. Our data suggest that the three low molecular weight endochorion proteins undergo disulfide bond crosslinking prior to oviposition in A. aegypti eggs, and that they undergo further crosslinking through dityrosine or trityrosine formation by peroxidase-catalyzed reactions. Our data suggest that chorion peroxidase is primarily responsible for the irreversible insolubilization of the three major endochorion proteins after oviposition. The molecular mechanisms of chorion hardening are also discussed.